Excitotoxicity and oxidative stress have separately been implicated in neuronal degeneration in acute neurologic insults and in age-related neurodegenerative diseases such as Parkinson's disease, amyotrophic lateral sclerosis and Huntington's Disease. Recent evidence suggests a link between excitotoxicity and oxidative stress. The mechanisms linking excitotoxicity, oxidative stress and neuronal degeneration are, however, poorly understood. Using kainic acid (KA) induced neuronal degeneration of cerebellar granule cells (CGC) as an in vitro model, this application proposes to test the hypothesis that oxidative damage to cellular macromolecules occurs following KA receptor activation, that cellular defense mechanisms play an important role in KA-induced neuronal degeneration, and that KA-induced oxidative stress may trigger apoptosis. Aim 1 will determine whether KA induced neuronal degeneration is associated with oxidative damage to DNA and proteins, using high performance liquid chromatography (HPLC) to measure 8- hydroxydeoxyguanosine and 2,4-dinitrophenylhydrazine spectrophotometry to measure protein carbonyl groups. Aim 2 will determine if reductions in free radical scavenging enzyme activities contribute to KA induced oxidative stress, using enzyme assays, pharmacologic agents, antisense phosphorothioate oligonucleotides, and transgenic mice. Aim 3 will test the hypothesis that oxidative stress induced by KA triggers apoptosis by evaluating chromatin condensation and DNA fragmentation and the effect of endonuclease, protein and RNA synthesis inhibition in KA toxicity. Successful completion of the proposed experiments will provide insight into the mechanisms of neuronal degeneration induced by excitatory amino acids and by free radicals. The results may suggest new avenues for prevention of neuronal death in stroke and in age-related neurodegenerative diseases.